The exchange of DNA segments by general and site-specific recombination is central to many biological processes. Recombination regulates the expression of specific genes, mediates viral integration and excision, and generates genetic diversity. It is also critical for the repair of DNA and transposition. REcombination requires the pairing of distinct sequences, breaking of DNA strands, and religation. The long-range goal of this project is to understand the mechanisms of these processes. In studies of site-specific recombination in vitro, the specific aims are to determine the topological parameters for five different enzymes, elucidate the role of supercoiling and the mechanism of enhancers for recombination, determine if DNA exchange is driven by protein subunit exchange, explore the structure of synaptic intermediates, and determine the order of strand breakage. In vivo, studies both in procaryotes and eucaryotes will focus on the establishment of controlled site-specific recombination systems such as Tn3 resolvase, phage lambda integrase, phage P1 Cre, and yeast Flp. These will permit us to contrast in vitro results with those in a variety of organisms and give valuable information on the state of supercoils in vivo. General recombination studies will concentrate on the structure of the paranemic joint formed by RecA protein and the establishment of defined in vivo and in vitro recombination systems. The methods involve enzymology, genetics and DNA topology. The techniques include electron microscopy of DNA and nucleoprotein complexes, gel electrophoresis, protein purification, and construction of topologically defined DNA substrates. The results of the proposed experiments should reveal basic characteristics of DNA structure and DNA-proteins interactions.